- The goals of this proposal are to elucidate the metabolic and biochemical effects of ATP depletion that are responsible for the major functional consequences of sublethal injury in renal tubular epithelial cells. The applicants will focus on two functional consequences of sublethal injury: loss of cell-matrix adhesion with resultant detachment of viable cells from the basement membrane and loss of cell-cell adhesion with consequent impairment of tight junction function. They will examine the effects of sublethal injury on three components of the cell cytoskeleton that are known to play an important role in maintaining normal tight junction function and cell-matrix adhesion; i) The actin cytoskeleton ii) the adhesion plaque, and iii) the adherens junction. The hypotheses underlying this application are: i) That the metabolic consequences of ATP depletion lead to disorganization of the actin cytoskeleton which in turn results in structural changes and function impairment of the adherens junctions and adhesion plaques, ii) that loss of the actin cytoskeleton is responsible for these effects by causing dysregulation of tyrosine phosphorylation of actin-binding proteins present within the adherens junctions and adhesion plaques, and iii) that interventions that prevent the changes in tyrosine phosphorylation associated with ATP depletion will ameliorate the functional consequences of sublethal injury. The specific aims are to identify the metabolic consequences of ATP depletion that result in structural disorganization of the actin cytoskeleton, to examine the effects of ATP depletion on tyrosine phosphorylation of proteins comprising the adhesion plaque and adherens junction, and to examine the role of these altered phosphorylation events in the functional consequences of sublethal injury.